HPAE-PAD determination of carbohydrates in honey to evaluate samples for quality and adulteration

Significance of the topic

Honey is a natural sweetener whose quality and purity are essential for consumer safety and market value. Comprehensive profiling of honey sugars helps assess authenticity, classify floral origin, and detect adulteration by inexpensive sugar syrups. High-resolution carbohydrate analysis underpins quality control in food laboratories and regulatory compliance.

Objectives and study overview

This study aimed to develop and validate a robust high-performance anion-exchange chromatography method with pulsed amperometric detection (HPAE-PAD) for quantifying mono-, di-, and oligosaccharides in honey. Twelve commercial honeys were characterized for their sugar profiles, and the method’s ability to detect and quantify intentional adulteration with various sugar syrups was evaluated.

Methodology and instrumentation

The method employed a Thermo Scientific Dionex ICS-5000+ system equipped with pulsed amperometric detection using gold on PTFE disposable electrodes. Separation was achieved on a Thermo Scientific Dionex CarboPac PA210-Fast-4μm analytical column (150×4 mm) preceded by a guard column, with a potassium hydroxide gradient elution (30 mM to 100 mM). Key parameters included a flow rate of 0.8 mL/min, 10 μL injections, and a total run time of 45 min. Samples were diluted, filtered, and analyzed without derivatization.

Main results and discussion

The method resolved 15 honey sugars—including glucose, fructose, nine disaccharides, and four trisaccharides—within 25 min. Calibration curves for 13 standards exhibited excellent linearity (r²>0.999). Monosaccharide content in honeys ranged from 63.7 to 81.4 g/100 g (fructose-to-glucose ratios between 1.01 and 1.37), meeting Codex standards. Disaccharides (e.g., maltose, sucrose) and trisaccharides (e.g., erlose, melezitose) varied with floral source. Spike recovery studies (n=4 honeys, two levels) demonstrated accuracies of 78–113%. Adulteration trials with corn, beet, and maple syrups at 10–20% levels showed clear shifts in sucrose, maltose, and sugar ratios, allowing detection of as little as 10% added syrup.

Benefits and practical applications

• Rapid, high-resolution profiling of honey sugars without derivatization
• Quantitative monitoring of key quality markers (reducing sugars, sucrose)
• Sensitive detection of multidimensional adulteration patterns
• Automated eluent generation reduces manual preparation and variability
• Applicability to routine quality control and authenticity testing in food industry laboratories

Future trends and opportunities

The integration of HPAE-PAD with chemometric analysis can further enhance floral classification and low-level adulteration detection. Advances in column chemistries may shorten run times and improve isomer resolution (e.g., turanose vs. palatinose). Online sample preparation and miniaturized systems could enable high-throughput screening. Adoption in regulatory frameworks and industry standards will drive broader application.

Conclusion

The validated HPAE-PAD method on CarboPac PA210-Fast-4μm columns provides a reliable, fast, and sensitive approach for comprehensive sugar profiling of honey. It meets international standards, delivers accurate quantification across a wide concentration range, and sensitively detects adulteration by commercial sugar syrups. This workflow supports robust quality control and authenticity assurance in honey production and trade.

References

1. Escuredo O, Dobre I, Fernández-González MC, Seijo MC. Contribution of botanical origin and sugar composition on honey crystallization. Food Chem. 2014;149:84–90.
2. Persano Oddo L, Piazza MG, Sabatini AG, Accorti M. Characterization of unifloral honeys. Apidologie. 1995;26:453–465.
3. Low NH, Sporns P. Analysis and quantitation of di- and trisaccharides in honey using capillary gas chromatography. J Food Sci. 1988;53:558–561.
4. Swallow KW, Low NH. Analysis and quantitation of carbohydrates in honey by HPLC. J Agric Food Chem. 1990;38(9):1828–1832.
5. Puscas A, Hosu A, Cimpoiu C. Thin-layer chromatographic method to control honey adulteration. J Chromatogr A. 2013;1272:132–135.
6. Ozcan M, Arslan D, Durmus AC. Effect of inverted saccharose on honey properties. Food Chem. 2006;99:24–29.
7. Codex Alimentarius Commission. Codex Standard for Honey. Rev. 2001;Std 12-1981.
8. Morales V, Corzo N, Sanz ML. HPAEC-PAD oligosaccharide analysis to detect honey adulteration. Food Chem. 2008;107:922–928.
9. Swallow KW, Low NH. Determination of honey authenticity by HPAEC-PAD. J AOAC Int. 1994;77(3):695–702.
10. Gasić E, et al. Phytochemical fingerprints of lime honey collected in Serbia. J AOAC Int. 2014;97(5):1259–1267.
11. Cordella CB, et al. Detection of honey adulteration via HPAEC-PAD and chemometrics. Anal Chim Acta. 2005;531:239–248.
12. Cordella CB, et al. Honey characterization and adulteration detection by pattern recognition on HPAEC-PAD profiles. J Agric Food Chem. 2003;51:3234–3242.
13. Thermo Fisher Scientific. Dionex CarboPac PA210-Fast-4µm Column Manual. 2016.
14. Thermo Fisher Scientific. CAN-123: Sugars in Honey Using HPAE-PAD. 2016.
15. White JW, Doner LW. Honey composition and properties. USDA Agric Handb. 1980;335:82–91.
16. Bogdanov S, et al. Honey for nutrition and health: a review. J Am Coll Nutr. 2008;27(6):677–689.
17. Ruiz-Matute AI, Brokl M, Soria AC, Sanz ML, Martínez-Castro I. GC-MS characterization of tri- and tetrasaccharides in honey. Food Chem. 2010;120:637–642.
18. Cotte JF, Casabianca H, Chardon S, et al. Chromatographic sugar analysis for monofloral honey characterization. Anal Bioanal Chem. 2004;380:698–705.